.alpha..sub.1 AT is a glycoprotein of approximately 53 kDA that functions as the major protease inhibitor in human serum. It is synthesized predominantly in the liver, although small quantities may also be produced locally, for example, by alveolar macrophages (White. R., D. Lee. G. G. Habicht and A. Janoff. 1981. Am. Rev. Respir. Dis. 123. 447). There has been much recent interest in the possible role of .alpha..sub.1 AT in the pathogenesis of many human diseases (reviewed in Lieberman. J.. 1980. in: Current Pulmonology, Vol. 2. ed. D. H. Simmond (Houghton. Mifflin Publishers. Boston, Mass.) pp. 41-68 and Fagerhol. M. K. and D. W. Cox. 1981. in: Advances in Human Genetics, Vol. 3. eds. H. Harris and K. Kirschhorn (Plenum Press. New York) pp. 1-62) because of its role in protecting tissues from damage resulting from local release of proteolytic enzymes and the demonstration that genetic variants of .alpha..sub.1 AT are associated with pulmonary and liver disease. Previously available methods for measuring .alpha..sub.1 AT in serum have proved less than satisfactory for measuring .alpha..sub.1 AT in other biologic fluids. For example, measurement by radial immunodiffusion (RID) frequently requires laborious concentration of fluids, with significant and unpredictable loss of .alpha..sub.1 AT (Warr. G. A., R. R. Martin, P. M. Sharp and R. D. Rossen. 1977, Am. Rev. Respir. Dis. 116, 25).
Methods previously used to quantitate .alpha..sub.1 AT include electroimmunodiffusion (Manildi. E. R., 1972. Clin. Chem. 18, 1019). nephelometry (Holzer, K. H. and G. Binzus, 1963. Verh. Dtsch. Ges. Inn. Med. 69. 456 and Gaidulis. L., H. A. Muensch, W. C. Maslow and W. Z. Borer, 1983, Clin. Chem. 29, 1838). and radial immunodiffusion (Mancini, G., A. O. Carbonara and J. F. Heremans. 1965, Int. J. Immunochem. 2, 235 and Dietz, A. A., H. M. Rubinstein and L. Hodges. 1974. Clin. Chem. 20. 396). All give comparable results in measuring serum .alpha..sub.1 AT. The simplicity of the RID method, and the commercial availability of RID kits for measuring .alpha..sub.1 AT, have made it the standard method of quantitating .alpha..sub.1 AT in serum. For the protective function of .alpha..sub.1 AT to be assessed, local concentrations should also be measured at sites of inflammation, protease release and tissue damage. For measuring .alpha..sub.1 AT in biologic fluids other than serum, the previously used methods have been less than satisfactory.
A number of investigators have measured .alpha..sub.1 AT in synovial fluids from patients with inflammatory diseases, where proteases are thought to play an important role in joint destruction (Swedlund H. A., G. G. Hunder and G. J. Gleich, 1974. Ann. Rheum. Dis. 33, 162: Brackertz, D., J. Hagmann and F. Kaepers. 1975, Ann. Rheum. Dis. 34. 225; Hadler, N. M., A. M. Johnson, J. K. Spitznagel and R. J. Quinet. 1981. Ann. Rheum. Dis. 40, 55: Ekerot. L. and K. Ohlsson. 1982. Rheumatol. Int. 2, 21: Pritchard. M. H., 1984. Ann. Rheum. Dis. 43, 50: and Virca, G. D., R. K. Mallya. M. Pepys and H. P. Schnebli, 1984. in: Proteases: Potential Role in Health and Disease, eds. W. H. Horl and A. Heidland (Plenum Press. New York)). Levels of .alpha..sub.1 AT in synovial fluids are roughly equivalent to serum levels, but investigators using different methods have obtained discrepant results. One reason appears to be that complexing of .alpha..sub.1 AT to proteases or to hyaluronic acid, both abundant in synovial fluid, impairs the migration of .alpha..sub.1 AT through gels, thus giving falsely low values (Molecular Biology of Human Proteins with Special Reference to Plasma Proteins, (Schultz. H. E. and J. F. Heremans eds., 1966. (Elsevier. New York)): Brackertz, D. J. Hagmann and F. Kaepers. 1975. Ann. Rheum. Dis. 34, 225; Ochi, T. K., K. Yonemasu and K. Ono, 1980. Ann. Rheum. Dis. 39. 235: Ekerot. L. and K. Ohlsson. 1982, Rheumatol. Int. 2. 21; and Cawston T. E., E. Mercer, M. DeSilva and B. L. Hazleman. 1984, Arthritis Rheum. 27, 285).
Measurement of .alpha..sub.1 AT in bronchoalveolar lavage fluids is of particular interest because of the association of .alpha..sub.1 AT deficiency with pulmonary disease. Several studies have attempted to measure .alpha..sub.1 AT in lavage fluids, and have found it necessary to concentrate the lavage fluid by membrane filtration before electroimmunodiffusion (Warr. G. A., R. R. Martin, P. M. Sharp and R. D. Rossen. 1977. Am. Rev. Respir. Dis. 116, 25.; Gadek, J. E., H. G. Klein, P. V. Holland and R. G. Crystal, 1981. J. Clin. Invest. 68. 1158) or RID quantitation (Olsen, G. N., J. O. Harris, J. R. Castle, R. H. Welchman and H. J. Karmgard, 1975, J. Clin. Invest. 55, 427: Niederman, M. S., L. L. Fritts, W. M. Merrill, R. B. Fick, R. A. Matthay, H. Y. Reynolds and J. B. L. Gee, 1984, Am. Rev. Respir. Dis. 129. 943). Concentrating lavage fluids resulted in the loss of 55%.+-.10% of the .alpha..sub.1 AT from the solutions, decreasing the accuracy of the results (Warr, G. A., R. R. Martin, P. M. Sharp and R. D. Rossen, 1977. Am. Rev. Respir. Dis. 116. 25).
Measurement of .alpha..sub.1 AT in human saliva has only recently been successful. Grimound and associates, using laser nephelometry, found a mean concentration of 0.57.+-.0.27 mg/dl of .alpha..sub.1 AT in saliva from 35 normal subjects (Grimound. A. M., D. Duffault, J. P. Lodter and J. P. Seguela, 1984, J. Biol. Buccale 12. 67). Two previous attempts to measure .alpha..sub.1 AT by electroimmunodiffusion found that levels measured were below the limits of detection by this technique (Ryley. H. C. and T. D. Brosan. 1973. J. Clin. Pathol. 26. 852; Gersel, A. V. and N. Pedersen, 1979, Int. J. Oral Surg. 8, 212).
So far as monoclonal antibodies to alpha-1-antitrypsin are concerned, only one is known. This is available from Cappel Laboratories (hereinafter referred to as the "Cappel antibody"). However, serious problems exist in use of the Cappel antibody in .alpha..sub.1 AT assays in that the Cappel antibody does not react strongly with .alpha..sub.1 AT and, even more importantly, the Cappel antibody cross reacts strongly with human albumin. The cross reaction with human albumin virtually eliminates the Cappel antibody as a candidate for a reagent to be used in any assay for .alpha..sub.1 AT employing body fluids.